<p>A novel optimised isolation method, TLC-bioautography, was evaluated and utilised in this research. Antibacterial compounds which were isolated from the dichloromethane extract of <i>Ferula ferulioides (Steud.) Korovin</i> were detected by means of the method. Their structures were elucidated by extensive spectral and chemical methods. Their antibacterial activities against drug-resistant <i>Staphylococcus aureus</i> (<i>S. aureus)</i> strains were evaluated with broth microdilution method, and the results proved that TLC-bioautography was an effective and highly efficient way to screen natural compounds from plant extracts against drug-resistant strains.</p>